The present invention relates to a method of detecting or quantifying a nucleic acid molecule present in a specimen.
Techniques for detecting a specific nucleic acid molecule present in a biological specimen are of importance in the fields of fundamental research and clinical study. The techniques are particularly important in analyzing proteins, which express and function at specific organs, at a nucleic-acid level. The techniques are also important in investigating expression of proteins and how to control the expression of proteins in the nervous system and the immune system associating signal transduction system. The techniques are extremely important in genetic diagnosis since mutant genes associated with genetic diseases, genes associated with cancer, and genes associated with viruses can be identified by these techniques.
A representative technique for detecting a nucleic acid molecule is a hybridization method. In this method, a target nucleic acid is hybridized with a nucleic acid probe having a complementary sequence of the target nucleic acid. Thereafter, the target nucleic acid can be identified using the probe. A drawback of this method resides in that it is difficult to detect a target nucleic acid if it is present in low numbers (e.g., about 1 to 1000 copies). Furthermore, the specificity is low. Therefore, it is difficult to recognize and detect analogous sequences distinguishably.
In a device called a DNA chip, various types of nucleic-acid probes are immobilized on a base. Detection using the detection method employing the DNA chip is much easier, compared to conventional hybridization methods. However, the probes immobilized on the base differ in optimal conditions, so that a false positive hybridization reaction may take place. This is a problem with this method. Furthermore, the design of the probe to be immobilized on the base must be changed in accordance with the nucleic acid to be detected. This means that an appropriate DNA chip must be prepared every time the nucleic acid to be detected is changed. Whereas, in clinical tests, various types of nucleic acids must be detected. Under these circumstances, conventionally a wide variety of DNA chips are required.
Conventional techniques, including the aforementioned methods, are complicated since an operation consists of many steps. In addition, a long detection time and a well-trained operator are required.
Gene diagnosis is usually used as a definite diagnosis. Therefore, a quick diagnosis is required without the possibility of miss-diagnosis. However, conventional methods do not fully satisfy these requirements.